Toxoplasma gondii is a common protozoan infection in man, with a sero-positivity in 30-50% of North Americans. Life-threatening toxoplasmosis occurs most often in patients with defective cell-mediated immunity who appear to reactivate previously acquired infection and develop severe encephalitis or disseminated disease. Life-threatening infections have occurred most often in organ-transplant recipients and patients with hematologic malignancies. More recently, toxoplasmosis has become a particularly prevalent opportunistic infection in patients with acquired immune deficiency syndrome (AIDS).
Systemic infection with toxoplasmosis and Pneumocistis carinii are generally treated with the antifolates pyrimethamine and trimethoprim, respectively. In the general population of patients with an intact immune system, these treatments have met with good success. However, over the past 3-4 years these infections have become widespread in the population of patients with AIDS. The AIDS syndrome develops following infection with a specific virus, HTLV-3, producing severe immune deficiency and increasing the risk of infection with opportunistic organisms such as Toxoplasmosis and P. carinii. Infection with these organisms in the AIDS population is extremely refractory to standard therapy, and many of those infected ultimately die from infectious complications. Therapy of Pneumocistis carinii pneumonia in the expanding population with AIDS has become problematic because of the high frequency of therapeutic failures. Because of the fastidious nature of the causative organisms, current treatment choices have been made with little information on the effects of the agents on the PC metabolic pathway.
Another problem encountered in treating patients with AIDS is the considerable number of patients (up to 60%) who develop allergic reactions or serious side effects during therapy with a sulfonamide and trimethoprim or pyrimethamine. No alternative regimen to sulfonamide and pyrimethamine has been found effective for the therapy of toxoplasma encephalitis.
Recently, the surge in the number of AIDS patients with toxoplasma encephalitis has highlighted the need for more effective therapies and for alternative drugs for those unable to tolerate pyrimethamine and sulfa drugs. Many AIDS patients develop life-threatening leukopenia, thrombocytopenia, hepatitis. or rash while receiving pyrimethamine and sulfa, yet no alternative regimen using these drugs singly or in combination with other drugs has been found to be effective.
Pneumocistis carinii (PC) pneumonia is the leading cause of death in the expanding population of patients with acquired immunodeficiency syndrome (AIDS). Current therapy for PC pneumonia consists of a dihydrofolate reductase inhibitor, most commonly the diaminopyrimidine trimethoprim, coupled with sulfa . . . oxazol, a dihydropteroate synthesis inhibitor. Presumably the synergistic interaction of these two compounds in the PC organism leads to folate depletion with resultant cessation of purine and thymidine synthesis and ultimately cell death.
Because of clinical resistance or drug intolerance, including allergic reactions and bone marrow suppression, approximately 40-60% of patients cannot be successfully treated. Because of the fastidious nature of this organism and its inability to grow in vitro, inhibition studies on key enzymes have been lacking. The choice of metabolic inhibitors used to treat PC pneumonia has been largely based on empiric data and the assumption that the DHFR in the causative organism is similar to bacteria and other protozoa in its sensitivity to the diaminopyrimidine inhibitors. While these inhibitors are apparently able to penetrate the cellular membrane of protozoa, there exists no data to allow a choice among the myriad available DHFR inhibitors. In contrast to these agents, highly potent DHFR inhibitors that retain the classic puridine structure such as methotrexate are ineffective in the treatment of bacterial and protozoal infections due to the lack of a specific membrane carrier required for the transport of these structures.
Dihydrofolate reductase (E.C.1.5.1.3;5,6,7,8-tetrahydrofolate:NADP+ oxoreductase) is required to maintain the intracellular pool of reduced folates in rapidly dividing cells. Inhibitors of this enzyme have proven effective in both antineoplastic and antimicrobial chemotherapy. Methotrexate, an analog that preserves the basic folate (pteroylglutamate) structure, potently inhibits dihyroreductase from mammalian and bacterial sources, but requires transport by a folate-specific membrane carrier found only on mammalian cells, and is therefore primarily useful as an antineoplastic agent. The diaminopyrimidines pyrimethamine and trimethaprim readily penetrate mammalian and microbial cells by diffusion, have intermediate inhibitory activity against bacterial dihydrofolate reductase but lesser potency against mammalian dihydrofolate reductase, and are used primarily in antibacterial and antiprotozoal therapy in combination with an inhibitor of folate synthesis such as sulfamethoxazole. While the diaminopyrimidines in combination with sulfonamides have become the primary form of therapy for certain parasitic infections, including toxoplasmosis, little is known concerning the transport of the various types of antifolates and the potency of their inhibition of dihydrofolate reductase in these organisms.
There is no current biochemical data addressing the ability of the currently used antifolates to inhibit the target enzyme, dihydrofolate reductase, in either toxoplasmosis or P. Carinii. Great variations exist in the ability of a given antifolate to inhibit the dihydrofolate reductase from different species. It has been speculated that pyrimethamine and trimethoprim are effective inhibitors of dihydrofolate reductase from these two organisms in a fashion similar to that observed in bacteria and plasmodia wherein the drugs are highly effective.